PCSK9 Locus Gene Editing With Donor Vectors for Stable Transgene Insertion

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Solution Overview

Problem

Current gene editing technologies lack effective methods for precisely targeting and modifying the PCSK9 gene locus for therapeutic interventions in genetic disorders, particularly in adult patients, and there is a need for improved compositions and methods to achieve stable, long-term therapeutic effects.

Innovation Solution

A dual component system comprising a gene editing vector and a donor vector, utilizing nucleases such as Cas9 or meganucleases, specifically targets the PCSK9 gene locus, enabling homology-directed recombination to insert and express exogenous transgenes, such as FIX, OTC, PAH, or LDLR, using AAV vectors with HDR arms and regulatory sequences to direct expression in target cells.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If nucleases are used to generate double strand breaks in the PCSK9 gene, then precise gene targeting is achieved, but DNA damage and potential off-target effects occur

Engineering Contradiction:

Improvegene targeting precisionVSAvoidDNA damage and off-target effects

Core Design Contradiction:

Measurement precisionVSObject-affected harmful factors

Solution Approach 1:

The patent converts the harmful double-strand breaks into beneficial homology-directed repair events by providing donor templates with homology arms that guide precise insertion of therapeutic transgenes at the PCSK9 locus, transforming DNA damage into a controlled gene editing mechanism

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

Solution Approach 2:

The patent introduces donor DNA templates as intermediary molecules that mediate between the nuclease-induced breaks and the desired therapeutic gene insertion, enabling precise targeting through homology-directed repair while controlling the editing outcome

Inventive Principle:

Principle #24Intermediary (Mediator)

2Manufacturing precision

If homology directed repair is used to insert transgenes, then precise genetic modification is achieved, but the process is inefficient and transient

Engineering Contradiction:

Improvegenetic modification precisionVSAvoidediting efficiency and durability

Core Design Contradiction:

Manufacturing precisionVSProductivity

Solution Approach 1:

The patent performs preliminary actions by pre-designing and providing donor templates with optimized homology arms and therapeutic transgenes before the editing process, enabling efficient and durable integration when the nuclease creates the double-strand break

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent changes parameters by optimizing the homology arm length, transgene sequence, and nuclease timing to maximize HDR efficiency and achieve stable, long-term expression of therapeutic genes in the PCSK9 locus

Inventive Principle:

Principle #35Parameter changes

3Measurement precision

If existing gene editing methods are used, then some level of gene targeting is achieved, but stable long-term therapeutic effects are not obtained

Engineering Contradiction:

Improvegene targeting capabilityVSAvoidtherapeutic effect duration

Core Design Contradiction:

Measurement precisionVSDuration of action of stationary object

Solution Approach 1:

The patent enables self-service by designing the editing system to leverage the cell's own homology-directed repair machinery, which automatically integrates the therapeutic transgene into the genome and maintains it long-term without external intervention

Inventive Principle:

Principle #25Self-service

Solution Approach 2:

The patent performs preliminary action by providing pre-designed donor templates with therapeutic transgenes that are ready for immediate integration upon nuclease-induced breaks, ensuring long-term expression through stable genomic integration rather than transient expression

Inventive Principle:

Principle #10Preliminary action

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

This approach allows for precise gene editing, reducing native PCSK9 expression and inserting therapeutic transgenes, providing stable, long-term therapeutic effects in treating genetic disorders like hemophilia B and liver metabolic disorders.

Implementation Method 1

Site-specific nucleases (such as CRISPR-Cas9 or meganucleases) generate double strand breaks (DSBs) in the chromosome, leading to DNA repair

Methodology Applied

Scientific EffectDouble strand break:

Implementation Method 2

In the presence of donor DNA, homology directed repair (HDR) occurs and replaces genetic information in the chromosome with new information from the donor gene

Methodology Applied

Scientific EffectHomology directed repair:

Data Source

PatentUS20260077061A1Compositions and methods for in vivo nuclease-mediated gene targeting for the treatment of genetic disorders in adult patients

Publication Date: 2026.03.19 THE TRUSTEES OF THE UNIV OF PENNSYLVANIA

AI summary

A dual component system for treating a genetic disorder is provided. The system includes (a) a gene editing vector comprising an expression cassette comprising a nucleic acid sequence encoding a nuclease and regulatory sequences that direct expression of the nuclease in a target cell comprising a PCSK9 gene; and (b) a donor vector comprising a nucleic acid sequence encoding an exogenous product for expression from the PCSK9 locus, wherein the inserted nucleic acid sequence does not encode PCSK9, wherein the system further comprises sequences that direct the nuclease to specifically targets the native PCSK9 gene locus; and wherein the native PCSK9 in the target cell is optionally ablated or reduced post-dosing with the dual component system.